We have established three cellular markers to distinguish between human noninvasive benign colon adenomas and invasive colon carcinomas in primary culture. The biologically relevant tumor cells are assayed directly from resected tumors. Carcinoma cells are primary cultured from resected tumors free of lymphocytes, endothelial cells, and fibroblasts, and remain viable and dividing in tissue culture for 2-3 weeks. This relative longevity allows the carcinoma cells to be modulated in tissue culture, and transition to a more benign state measured by the markers developed. In initial experiments, the differentiation agent, hexamethylene bisacetamide (HMBA), induced differentiation of three tumors graded as moderately differentiated by histopathology; HMBA was ineffective on one poorly differentiated and two colloid colon carcinomas. We propose (1) to study the response of 20 primary cultured human colon carcinomas to HMBA by our in vitro assays; (2) to compare the effects of HMBA and TGFbeta on primary-cultured carcinomas; (3) to assay for terminal differentiation of normal colonic epithelial cells by TGFbeta; (4) to test the growth potential of the HMBA-treated cells both in vivo in the subrenal capsule assay in nude mice and in vitro by assay of the tumor growth fraction by continuous labeling with 3H-thymidine, (5) to begin studies to purify factor(s) secreted by HMBA-differentiated HT29 cells.